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Engineering Escherichia coli for Poly-β-hydroxybutyrate Production from Methanol

The naturally occurring one-carbon assimilation pathways for the production of acetyl-CoA and its derivatives often have low product yields because of carbon loss as CO(2). We constructed a methanol assimilation pathway to produce poly-3-hydroxybutyrate (P3HB) using the MCC pathway, which included t...

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Autores principales: Wang, Jiaying, Chen, Zhiqiang, Deng, Xiaogui, Yuan, Qianqian, Ma, Hongwu
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10135841/
https://www.ncbi.nlm.nih.gov/pubmed/37106602
http://dx.doi.org/10.3390/bioengineering10040415
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author Wang, Jiaying
Chen, Zhiqiang
Deng, Xiaogui
Yuan, Qianqian
Ma, Hongwu
author_facet Wang, Jiaying
Chen, Zhiqiang
Deng, Xiaogui
Yuan, Qianqian
Ma, Hongwu
author_sort Wang, Jiaying
collection PubMed
description The naturally occurring one-carbon assimilation pathways for the production of acetyl-CoA and its derivatives often have low product yields because of carbon loss as CO(2). We constructed a methanol assimilation pathway to produce poly-3-hydroxybutyrate (P3HB) using the MCC pathway, which included the ribulose monophosphate (RuMP) pathway for methanol assimilation and non-oxidative glycolysis (NOG) for acetyl-CoA (precursor for PHB synthesis) production. The theoretical product carbon yield of the new pathway is 100%, hence no carbon loss. We constructed this pathway in E. coli JM109 by introducing methanol dehydrogenase (Mdh), a fused Hps–phi (hexulose-6-phosphate synthase and 3-phospho-6-hexuloisomerase), phosphoketolase, and the genes for PHB synthesis. We also knocked out the frmA gene (encoding formaldehyde dehydrogenase) to prevent the dehydrogenation of formaldehyde to formate. Mdh is the primary rate-limiting enzyme in methanol uptake; thus, we compared the activities of three Mdhs in vitro and in vivo and then selected the one from Bacillus methanolicus MGA3 for further study. Experimental results indicate that, in agreement with the computational analysis results, the introduction of the NOG pathway is essential for improving PHB production (65% increase in PHB concentration, up to 6.19% of dry cell weight). We demonstrated that PHB can be produced from methanol via metabolic engineering, which provides the foundation for the future large-scale use of one-carbon compounds for biopolymer production.
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spelling pubmed-101358412023-04-28 Engineering Escherichia coli for Poly-β-hydroxybutyrate Production from Methanol Wang, Jiaying Chen, Zhiqiang Deng, Xiaogui Yuan, Qianqian Ma, Hongwu Bioengineering (Basel) Article The naturally occurring one-carbon assimilation pathways for the production of acetyl-CoA and its derivatives often have low product yields because of carbon loss as CO(2). We constructed a methanol assimilation pathway to produce poly-3-hydroxybutyrate (P3HB) using the MCC pathway, which included the ribulose monophosphate (RuMP) pathway for methanol assimilation and non-oxidative glycolysis (NOG) for acetyl-CoA (precursor for PHB synthesis) production. The theoretical product carbon yield of the new pathway is 100%, hence no carbon loss. We constructed this pathway in E. coli JM109 by introducing methanol dehydrogenase (Mdh), a fused Hps–phi (hexulose-6-phosphate synthase and 3-phospho-6-hexuloisomerase), phosphoketolase, and the genes for PHB synthesis. We also knocked out the frmA gene (encoding formaldehyde dehydrogenase) to prevent the dehydrogenation of formaldehyde to formate. Mdh is the primary rate-limiting enzyme in methanol uptake; thus, we compared the activities of three Mdhs in vitro and in vivo and then selected the one from Bacillus methanolicus MGA3 for further study. Experimental results indicate that, in agreement with the computational analysis results, the introduction of the NOG pathway is essential for improving PHB production (65% increase in PHB concentration, up to 6.19% of dry cell weight). We demonstrated that PHB can be produced from methanol via metabolic engineering, which provides the foundation for the future large-scale use of one-carbon compounds for biopolymer production. MDPI 2023-03-26 /pmc/articles/PMC10135841/ /pubmed/37106602 http://dx.doi.org/10.3390/bioengineering10040415 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Wang, Jiaying
Chen, Zhiqiang
Deng, Xiaogui
Yuan, Qianqian
Ma, Hongwu
Engineering Escherichia coli for Poly-β-hydroxybutyrate Production from Methanol
title Engineering Escherichia coli for Poly-β-hydroxybutyrate Production from Methanol
title_full Engineering Escherichia coli for Poly-β-hydroxybutyrate Production from Methanol
title_fullStr Engineering Escherichia coli for Poly-β-hydroxybutyrate Production from Methanol
title_full_unstemmed Engineering Escherichia coli for Poly-β-hydroxybutyrate Production from Methanol
title_short Engineering Escherichia coli for Poly-β-hydroxybutyrate Production from Methanol
title_sort engineering escherichia coli for poly-β-hydroxybutyrate production from methanol
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10135841/
https://www.ncbi.nlm.nih.gov/pubmed/37106602
http://dx.doi.org/10.3390/bioengineering10040415
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